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Proceedings of the National Academy of... Oct 2023Most biocatalytic processes in eukaryotic cells are regulated by subcellular microenvironments such as membrane-bound or membraneless organelles. These natural...
Most biocatalytic processes in eukaryotic cells are regulated by subcellular microenvironments such as membrane-bound or membraneless organelles. These natural compartmentalization systems have inspired the design of synthetic compartments composed of a variety of building blocks. Recently, the emerging field of liquid-liquid phase separation has facilitated the design of biomolecular condensates composed of proteins and nucleic acids, with controllable properties including polarity, diffusivity, surface tension, and encapsulation efficiency. However, utilizing phase-separated condensates as optical sensors has not yet been attempted. Here, we were inspired by the biosynthesis of melanin pigments, a key biocatalytic process that is regulated by compartmentalization in organelles, to design minimalistic biomolecular condensates with emergent optical properties. Melanins are ubiquitous pigment materials with a range of functionalities including photoprotection, coloration, and free radical scavenging activity. Their biosynthesis in the confined melanosomes involves oxidation-polymerization of tyrosine (Tyr), catalyzed by the enzyme tyrosinase. We have now developed condensates that are formed by an interaction between a Tyr-containing peptide and RNA and can serve as both microreactors and substrates for tyrosinase. Importantly, partitioning of Tyr into the condensates and subsequent oxidation-polymerization gives rise to unique optical properties including far-red fluorescence. We now demonstrate that individual condensates can serve as sensors to detect tyrosinase activity, with a limit of detection similar to that of synthetic fluorescent probes. This approach opens opportunities to utilize designer biomolecular condensates as diagnostic tools for various disorders involving abnormal enzymatic activity.
Topics: RNA; Melanins; Monophenol Monooxygenase; Proteins; Peptides; Organelles
PubMed: 37871222
DOI: 10.1073/pnas.2310569120 -
Experimental Dermatology Jul 2017In living cells, melanin pigment is formed within melanosomes, which not only protect the cells from autodestruction, but also serve as second messenger organelles...
In living cells, melanin pigment is formed within melanosomes, which not only protect the cells from autodestruction, but also serve as second messenger organelles regulating important skin functions, with melanocytes acting as primary sensory and regulatory cells of the epidermis. Yet, one can argue that skin melanin, which may negatively affect cellular homeostasis in melanoma, really exerts protective functions. Consequently, the actual functions of melanin and the melanogenic pathway in skin biology remains enigmatic. Yet, the solution of this riddle seems simple - to check the actual influence of natural melanin on skin cells in the dark. Since many interesting hypotheses and theories put forward in this respect did not survive confrontation with the experiment, a leading pigment research group from Naples was brave to "jump off the cliff" by confronting theory with experimental reality. They showed that, in the dark, human hair-derived melanin promotes inflammatory responses in keratinocytes, lowers their viability, promotes oxidative stress, and that pheomelanin does so more strongly than eumelanin. Thus, pheomelanin hardly protects red-haired individuals, even when avoiding the sun. Black hairs do not do much better either, unless they undergo graying.
Topics: Cell Survival; Darkness; Epidermis; Hair Color; Humans; Inflammation; Keratinocytes; Light; Melanins; Melanocytes; Melanosomes; Oxidative Stress; Skin; Skin Pigmentation
PubMed: 27541811
DOI: 10.1111/exd.13171 -
The Journal of Investigative Dermatology Feb 2023Palmitoylation is a lipid modification involving the attachment of palmitic acid to a cysteine residue, thereby affecting protein function. We investigated the effect of...
Palmitoylation is a lipid modification involving the attachment of palmitic acid to a cysteine residue, thereby affecting protein function. We investigated the effect of palmitoylation of tyrosinase, the rate-limiting enzyme in melanin synthesis, using a human three-dimensional skin model system and melanocyte culture. The palmitoylation inhibitor, 2-bromopalmitate, increased melanin content and tyrosinase protein levels in melanogenic cells by suppressing tyrosinase degradation. The palmitoylation site was Cysteine in the C-terminal cytoplasmic tail of tyrosinase. The nonpalmitoylatable mutant, tyrosinase (C500A), was slowly degraded and less ubiquitinated than wild-type tyrosinase. Screening for the Asp-His-His-Cys (DHHC) family of proteins for tyrosinase palmitoylation suggested that DHHC2, 3, 7, and 15 are involved in tyrosinase palmitoylation. Knockdown of DHHC2, 3, or 15 increased tyrosinase protein levels and melanin content. Determination of their subcellular localization in primary melanocytes revealed that DHHC2, 3, and 15 were localized in the endoplasmic reticulum, Golgi apparatus, and/or melanosomes, whereas only DHHC2 was localized in the melanosomes. Immunoprecipitation showed that DHHC2 and DHHC3 predominantly bind to mature and immature tyrosinase, respectively. Taken together, tyrosinase palmitoylation at Cysteine by DHHC2, 3, and/or 15, especially DHHC2 in trans-Golgi apparatus and melanosomes and DHHC3 in the endoplasmic reticulum and cis-Golgi apparatus, regulate melanogenesis by modulating tyrosinase protein levels.
Topics: Humans; Monophenol Monooxygenase; Cysteine; Lipoylation; Acyltransferases; Melanins; Melanocytes
PubMed: 36063887
DOI: 10.1016/j.jid.2022.08.040 -
Magnetic Resonance in Medicine Nov 2017To investigate the physical mechanisms associated with the contrast observed in neuromelanin MRI.
PURPOSE
To investigate the physical mechanisms associated with the contrast observed in neuromelanin MRI.
METHODS
Phantoms having different concentrations of synthetic melanins with different degrees of iron loading were examined on a 3 Tesla scanner using relaxometry and quantitative magnetization transfer (MT).
RESULTS
Concentration-dependent T and T shortening was most pronounced for the melanin pigment when combined with iron. Metal-free melanin had a negligible effect on the magnetization transfer spectra. On the contrary, the presence of iron-laden melanins resulted in a decreased magnetization transfer ratio. The presence of melanin or iron (or both) did not have a significant effect on the macromolecular content, represented by the pool size ratio.
CONCLUSION
The primary mechanism underlying contrast in neuromelanin-MRI appears to be the T reduction associated with melanin-iron complexes. The macromolecular content is not significantly influenced by the presence of melanin with or without iron, and thus the MT is not directly affected. However, as T plays a role in determining the MT-weighted signal, the magnetization transfer ratio is reduced in the presence of melanin-iron complexes. Magn Reson Med 78:1790-1800, 2017. © 2016 International Society for Magnetic Resonance in Medicine.
Topics: Humans; Iron; Magnetic Resonance Imaging; Melanins; Models, Biological; Phantoms, Imaging; Substantia Nigra
PubMed: 28019018
DOI: 10.1002/mrm.26584 -
Molecules (Basel, Switzerland) Apr 2021This study was performed to clarify the inhibitory effects of cycloheterophyllin on melanin synthesis. In order to elucidate the inhibitory effects of cycloheterophyllin...
This study was performed to clarify the inhibitory effects of cycloheterophyllin on melanin synthesis. In order to elucidate the inhibitory effects of cycloheterophyllin on the B16F10 cell line, cell viability, messenger ribonucleic acid (mRNA) expressions, tyrosinase activity assay, and melanin production assay were measured. The effects of cycloheterophyllin on tyrosinase-related protein 1 ()//tyrosinase ()/microphthalmia-associated transcription factor () mRNA expressions and melanin content were determined. Quantitative real-time RT-PCR showed that cycloheterophyllin decreased the mRNA expression level of genes and melanin production contents than α-MSH-treated B16F10 cells. The tyrosinase activity assay revealed that cycloheterophyllin decreased the melanin production in the B16F10 cells. These data show that cycloheterophyllin increases the whitening effects in the B16F10 cells; thus, cycloheterophyllin is a potent ingredient for skin whitening. Thus, further research on the mechanism of action of cycloheterophyllin for the development of functional materials should be investigated.
Topics: Animals; Antineoplastic Agents; Antioxidants; Biosynthetic Pathways; Dose-Response Relationship, Drug; Flavonoids; Gene Expression Regulation, Enzymologic; Gene Expression Regulation, Neoplastic; Humans; Melanins; Melanoma, Experimental; Mice
PubMed: 33926006
DOI: 10.3390/molecules26092526 -
Optics Express Aug 2022Pump-probe microscopy of melanin in tumors has been proposed to improve diagnosis of malignant melanoma, based on the hypothesis that aggressive cancers disaggregate...
Pump-probe microscopy of melanin in tumors has been proposed to improve diagnosis of malignant melanoma, based on the hypothesis that aggressive cancers disaggregate melanin structure. However, measured signals of melanin are complex superpositions of multiple nonlinear processes, which makes interpretation challenging. Polarization control during measurement and data fitting are used to decompose signals of melanin into their underlying molecular mechanisms. We then identify the molecular mechanisms that are most susceptible to melanin disaggregation and derive false-coloring schemes to highlight these processes in biological tissue. We demonstrate that false-colored images of a small set of melanoma tumors correlate with clinical concern. More generally, our systematic approach of decomposing pump-probe signals can be applied to a multitude of different samples.
Topics: Humans; Melanins; Melanoma; Microscopy; Skin Neoplasms
PubMed: 36242259
DOI: 10.1364/OE.469506 -
MBio Jun 2022Contamination of food and feed with toxin-producing fungi is a major threat in agriculture and for human health. The filamentous fungus Alternaria alternata is one of...
Contamination of food and feed with toxin-producing fungi is a major threat in agriculture and for human health. The filamentous fungus Alternaria alternata is one of the most widespread postharvest contaminants and a weak plant pathogen. It produces a large variety of secondary metabolites with alternariol and its derivatives as characteristic mycotoxin. Other important phyto- and mycotoxins are perylene quinones (PQs), some of which have anticancer properties. Here, we discovered that the PQ altertoxin (ATX) biosynthesis shares most enzymes with the 1,8-dihydroxynaphthalene (1,8-DHN) melanin pathway. However, melanin was formed in aerial hyphae and spores, and ATXs were synthesized in substrate hyphae. This spatial separation is achieved through the promiscuity of a polyketide synthase, presumably producing a pentaketide (T4HN), a hexaketide (AT4HN), and a heptaketide (YWA1) as products. T4HN directly enters the altertoxin and DHN melanin pathway, whereas AT4HN and YWA1 can be converted only in aerial hyphae, which probably leads to a higher T4HN concentration, favoring 1,8-DHN melanin formation. Whereas the production of ATXs was strictly dependent on the CmrA transcription factor, melanin could still be produced in the absence of CmrA to some extent. This suggests that different cues regulate melanin and toxin formation. Since DHN melanin is produced by many fungi, PQs or related compounds may be produced in many more fungi than so far assumed. Mycotoxins are a major threat for human health. Food safety control relies on the identification of the toxins or the detection of the expression of the respective genes. The latter method, however, relies on the knowledge of the biosynthetic pathway and the key genes. Alternaria alternata is a major food contaminant and produces many different mycotoxins with altertoxins and other perylene quinones as prominent examples. Here, we discovered that the biosynthetic pathway for altertoxins shares most of the enzymes with the dihydroxynaphthalene (DHN) melanin pathway. Because the DHN melanin pathway is widespread among fungi, the production of mycotoxins of the perylene quinone class could be more widespread than so far anticipated.
Topics: Alternaria; Humans; Melanins; Mycotoxins; Perylene; Quinones
PubMed: 35475649
DOI: 10.1128/mbio.00219-22 -
Experimental Eye Research Sep 2014The retinal pigment epithelium contains three major types of pigment granules; melanosomes, lipofuscin and melanolipofuscin. Melanosomes in the retinal pigment... (Review)
Review
The retinal pigment epithelium contains three major types of pigment granules; melanosomes, lipofuscin and melanolipofuscin. Melanosomes in the retinal pigment epithelium (RPE) are formed during embryogenesis and mature during early postnatal life while lipofuscin and melanolipofuscin granules accumulate as a function of age. The difficulty in studying the formation and consequences of melanosomes and lipofuscin granules in RPE cell culture is compounded by the fact that these pigment granules do not normally occur in established RPE cell lines and pigment granules are rapidly lost in adult human primary culture. This review will consider options available for overcoming these limitations and permitting the study of melanosomes and lipofuscin in cell culture and will briefly evaluate the advantages and disadvantages of the different protocols.
Topics: Animals; Cells, Cultured; Epithelial Cells; Humans; Lipofuscin; Melanins; Models, Animal; Models, Biological; Retinal Pigment Epithelium
PubMed: 25152361
DOI: 10.1016/j.exer.2014.01.016 -
Cells May 2023Based on traditional pharmacological applications and partial in vitro data, (CA) is proposed to act on skin whitening. However, its functional evaluation and...
Based on traditional pharmacological applications and partial in vitro data, (CA) is proposed to act on skin whitening. However, its functional evaluation and underlying mechanisms have yet to be identified. This study aimed to examine the anti-melanogenesis activity of CA fraction B (CAFB) on UVB-induced skin hyperpigmentation. Forty C57BL/6j mice were exposed to UVB (100 mJ/cm, five times/week) for eight weeks. After irradiation, CAFB was applied to the left ear once a day for 8 weeks (the right ear served as an internal control). The results showed that CAFB significantly reduced melanin production in the ear skin, as indicated by the gray value and Mexameter melanin index. In addition, CAFB treatment notably decreased melanin production in α-MSH-stimulated B16F10 melanocytes, along with a significant reduction in tyrosinase activity. Cellular cAMP (cyclic adenosine monophosphate), MITF (microphthalmia-associated transcription factor), and tyrosinase-related protein 1 (TRP1) were also noticeably downregulated by CAFB. In conclusion, CAFB is a promising ingredient for treating skin disorders caused by the overproduction of melanin and its underlying mechanisms involving the modulation of tyrosinase, mainly mediated by the regulation of the cAMP cascade and MITF pathway.
Topics: Animals; Mice; Melanins; Vincetoxicum; Monophenol Monooxygenase; Microphthalmia-Associated Transcription Factor; Signal Transduction; Mice, Inbred C57BL
PubMed: 37408224
DOI: 10.3390/cells12101390 -
Environmental Microbiology Reports Aug 2022As human activity in space continues to increase, understanding how biological assets respond to spaceflight conditions is becoming more important. Spaceflight...
As human activity in space continues to increase, understanding how biological assets respond to spaceflight conditions is becoming more important. Spaceflight conditions include exposure to ionizing radiation, microgravity, spacecraft vibrations and hypervelocity; all of which can affect the viability of biological organisms. Previous studies have shown that melanin-producing fungi are capable of surviving the vacuum of space and Mars-simulated conditions in Low Earth Orbit. This survival has been associated in part with the protective effects of melanin, but a comparison of fungal viability in the presence or absence of melanin following spaceflight has never been tested. In this study, we evaluated the protective effects of melanin by comparing the viability of melanized and non-melanized clones of Cryptococcus neoformans yeasts following a roundtrip to the International Space Station. Yeast colonies were placed inside two MixStix silicone tubes; one stayed on Earth and the other was transported inside for 29 days before returning to Earth. Post-flight analysis based on colony-forming unit numbers shows that melanized yeast viability was 50% higher than non-melanized yeasts, while no difference was observed between the Earth-bound control samples. The results suggest that fungal melanin could increase the lifespan of biological assets in space.
Topics: Cryptococcus neoformans; Humans; Melanins; Saccharomyces cerevisiae; Space Flight
PubMed: 35852045
DOI: 10.1111/1758-2229.13078